Differential expression of BAFF and its receptors in discoid lupus erythematosus patients

Abstract

Background: B-cell activating factor of the TNF family (BAFF) promotes the maturation and survival of B cells. Because BAFF levels are elevated in systemic lupus erythematosus (SLE) patients, BAFF has been the target of emerging therapies for SLE, such as belimumab. Levels of BAFF and its receptors in discoid lupus erythematosus (DLE) patients are unknown.

Objective: To compare skin and blood mRNA and protein levels of BAFF and its receptors BAFF-R, TACI, and BCMA in DLE subjects with (DLE+/SLE+ (N=28)) and without SLE (DLE+/SLE- (N=35)), psoriasis subjects (N=11), and normal subjects (N=42).

Methods: We used quantitative real-time PCR to measure blood and skin BAFF, BAFF-R, TACI, and BCMA mRNA, sandwich ELISAs to measure sera BAFF, and immunohistochemistry to evaluate BAFF and BAFF-R skin protein expression.

Results: BAFF mRNA and protein levels were highest in DLE+/SLE+blood, followed by DLE+/SLE-, psoriasis, and normal blood. BAFF protein also correlated with anti-nuclear antibodies, and autoantibodies against double-stranded DNA, single-stranded DNA, and ribonucleoprotein, and Systemic Lupus Erythematosus Disease Activity Index scores in DLE patients. While showing no difference between DLE+/SLE+ and DLE+/SLE- skin, BAFF and its receptors mRNA were up-regulated in DLE skin vs. normal and psoriasis skin. DLE skin had higher percentages of BAFF-R⁺ inflammatory cells, likely T cells and macrophages, than psoriasis and normal skin.

Conclusions: BAFF may be a serologic marker of systemic disease in DLE patients. BAFF and its receptors are elevated in DLE skin, suggesting that targeted therapies against these proteins could treat refractory DLE patients.

Keywords: BAFF; BAFF-R; Discoid lupus erythematosus; Macrophage; Systemic lupus erythematosus; T cell.

Fig. 1

mRNA levels of BAFF, but not its receptors, are elevated in DLE+/SLE+ blood versus DLE+/SLE−, psoriasis, and normal blood. Quantitative real-time PCR was performed to evaluate mRNA levels of BAFF (A), and its receptors, BAFF-R (B), TACI (C), and BCMA (D) in the whole blood of DLE+/SLE+ (N=27), DLE+/SLE− (N=28), psoriasis (N=11), and normal patients (N=17). We performed secondary analyses using Kruskal Wallis test. Capped bars between groups with astericks represent significant differences between groups based on Dunn’s test for multiple comparisons. *: p<0.05.

Fig. 2

BAFF protein levels are elevated in DLE+/SLE+ sera versus DLE+/SLE−, normal and blood sera and correlate with levels of BAFF mRNA, various autoantibodies, and SLEDAI scores. Sandwich ELISAs were performed to evaluate sera levels of BAFF in DLE+/SLE+ (N=28), DLE+/SLE− (N=35), psoriasis (N=11), and normal (N=32) patients (A). We performed secondary analyses using Kruskal Wallis test. Capped bars between groups with astericks represent significant differences between groups based on Dunn’s test for multiple comparisons. Protein (pg/mL) and mRNA levels (fold change) of BAFF in the sera of DLE+/SLE+ (N=27) (dark circles), DLE+/SLE− (N=28) (white circles), psoriasis (dark squares) (N=11), and normal (white squares) (N=17) patients were correlated (B). BAFF protein and levels of ANA (C), anti-dsDNA (D), anti-ssDNA (E), and anti-RNP (F) antibodies and SLEDAI scores (G) in the sera of 28 DLE+/SLE+ (dark circles) and 35 DLE+/SLE− (white circles) patients were correlated. Spearman’s correlation coefficients and corresponding p-values were calculated. *: p<0.05.

Fig. 2

BAFF protein levels are elevated in DLE+/SLE+ sera versus DLE+/SLE−, normal and blood sera and correlate with levels of BAFF mRNA, various autoantibodies, and SLEDAI scores. Sandwich ELISAs were performed to evaluate sera levels of BAFF in DLE+/SLE+ (N=28), DLE+/SLE− (N=35), psoriasis (N=11), and normal (N=32) patients (A). We performed secondary analyses using Kruskal Wallis test. Capped bars between groups with astericks represent significant differences between groups based on Dunn’s test for multiple comparisons. Protein (pg/mL) and mRNA levels (fold change) of BAFF in the sera of DLE+/SLE+ (N=27) (dark circles), DLE+/SLE− (N=28) (white circles), psoriasis (dark squares) (N=11), and normal (white squares) (N=17) patients were correlated (B). BAFF protein and levels of ANA (C), anti-dsDNA (D), anti-ssDNA (E), and anti-RNP (F) antibodies and SLEDAI scores (G) in the sera of 28 DLE+/SLE+ (dark circles) and 35 DLE+/SLE− (white circles) patients were correlated. Spearman’s correlation coefficients and corresponding p-values were calculated. *: p<0.05.

Fig. 3

mRNA levels of BAFF and its receptors are up-regulated in DLE skin versus psoriasis and normal skin. Quantitative real-time PCR was performed to evaluate mRNA levels of BAFF (A), and its receptors, BAFF-R (B), TACI (C), and BCMA (D) in the skin of DLE (N=20), psoriasis (N=5), and normal (N=10) patients. We performed secondary analyses using Kruskal Wallis test. Capped bars between groups with astericks represent significant differences between groups based on Dunn’s test for multiple comparisons. mRNA levels of BAFF in the skin and blood of DLE (N=19) and psoriasis (N=5) patients were correlated (E). Spearman’s correlation coefficient and corresponding p-value were calculated. *: p<0.05, **: p<0.005, ***: p<0.0005.

Fig. 4

BAFF-R is expressed by a greater percentage of inflammatory cells in DLE skin (N=14) versus psoriasis (N=4) and normal (N=6) skin. Immunohistochemical analysis of BAFF-R (brown) were performed on representative samples of DLE (A), psoriasis (B), and normal (C) skin. Magnification: 100×.

Fig. 5

BAFF and BAFF-R expression correspond to cells expressing CD3 and CD163 in DLE skin. Immunohistochemical stainings of BAFF (A), BAFF-R (B), T-cell marker CD3 (C), macrophage marker CD163 (D), and B-cell marker CD20 (E) were performed on another representative sample of DLE skin. Magnification: 100×.